Q: What are the best practices for conducting impurity profiling of drugs? What is the best approach to finding unknown impurities?
In order to safeguard the quality, safety, and efficacy of medicines, impurity profiling of drugs is paramount. The chemical
structure of these impurities is usually very much like that of the API and therefore the separation of API from impurities
can be a challenge. As a result, drug producers use methods with the highest possible resolution to study the related substances
present in a drug.
When aiming to find all the impurities in a drug, it is advisable to implement various kinds of separation techniques. For
instance, LC can have a different separation selectivity from capillary electrophoresis (CE) so applying both techniques yields
supplementary and complementary information about a particular sample. Moreover, within chromatography, it is often advisable
to use a combination of columns with different selectivities, that is, to apply orthogonal methods.
Another way of finding all the impurities in a sample is to combine a separation technique with a variety of detectors. Each
type of detector can highlight different types of compounds because the detector response depends on the chemical structure
of the compound. Does it have a UV chromophore? Does it exhibit good ionization in an MS probe? Does it show good conductivity?
The answers to these questions point to different detectors.
The procedure applied strongly depends on the level of knowledge of the drug substance, the phase of development, and the
purpose of the impurity profiling study. For well defined processes, applying the defined method of analysis for different
batches could be an acceptable profiling approach. However, state-of-the-art profiling would require a different approach,
starting with performing a theoretical assessment based on product knowledge and the literature on the synthesis, degradation
pathways, and interaction with excipients. Based on the long list of impurities from the assessment, selection of the analytical
techniques and methods will be made, taking into account the physical and chemical characteristics of the components, such
as presence of chromophores, (calculated) pKa values, and (predicted) ionization in MS. After tuning the methods based on
a set of representative components, the data are processed, applying peak picking software to enable any impurities to be
found or to compare (differences in) impurity levels. This approach, combining prior knowledge, good quality data, and suitable
software, enhances the chance of detecting unknowns and increases the understanding of impurities found. When the information
is added to a product knowledge document, it allows the information level to be monitored and estimations to be made at any
point in time.
It is not really possible to point out a single procedure that is routinely applicable for impurity profiling. The very diverse
nature of (potential) impurities demands significant expertise in the selection of separation and detection techniques to
be included in impurity profiling studies. A theoretical expert assessment of impurities that might be expected should therefore
always precede any effort of practical profiling. Based on the estimated properties of these potential impurities, a choice
can be made for an array of separation and detection techniques to include in such studies. These separation techniques should
preferably be selected so as to be orthogonal (that is, relying on significantly differing separation mechanisms; for example,
reversed-phase LC [various modes], hydrophilic interaction chromatography [HILIC], CE) to minimize the chances of missing
out on impurities that may potentially co-elute, elute without retention, or not elute at all. In addition, for detection,
it is desirable to include more than just a single technique (note that no single detection technique is really generic).
Finally, after profiling, investigation of mass balance proves to be a versatile tool to estimate if important impurities
may have been missed.
Q: What are important topics for research in pharmaceutical analysis of small molecules?
For small molecules, analysts always aim for an improvement in the sensitivity of the analysis and an improvement in selectivity
of the separation technique, as well as an improvement in efficiency and speed. This is why we will most probably witness
in the near future more and more methods using ultrahigh-pressure liquid chromatography (UHPLC), entering into monographs.
Companies can save a lot of time and expense by adopting the newer miniaturized separation techniques, and they have an advantage
in doing so from the start, that is, when submitting the regulatory file. They can avoid the time-consuming method transfer
and adjustment process needed to transfer a standard LC method to a miniaturized one.
In general, activities that support impurity profiling offer opportunities for improvement; for example, the improvement of
method development strategies, orthogonality of methods and techniques, column selection, prediction of degradation pathways,
and interaction with excipients. Analysis of polar components is of special interest as these show little retention in the
classic LC–UV methods on C18 columns; investigation into the use of HILIC separation methods have grown in the last few years
as a result. Control of potential genotoxic impurities is also an important area for research. In contrast to impurity profiling
of regular impurities, which focuses on the detection of any unknowns above a specific threshold, the control of potential
genotoxic impurities today fully relies on assessments. Although the impurity threshold for genotoxic impurities is much lower,
technical capabilities allow screening for toxic impurities based on their intrinsic reactivity, in addition to the assessment.
Although not required, these screening methods have already been developed for alkylation agents and a similar approach would
allow other classes of toxic compounds to be screened for. With new EMA (CHMP/SWP/4446/2000; 2013), USP (232/233; 2014), and
ICH (Q3D; 2013) guidance on the horizon for heavy metals, there is a strong increase in work performed in this field, mainly
using inductively coupled plasma–mass spectrometry (ICP–MS).
Important areas include: genotoxic impurities; residues of (heavy) metal (catalysts); and process analytical technology.